While plant rRNA gene organization and expression have been studied for several decades, the repetitive nature and high sequence identity between the tandemly-repeated units has prevented precise studies to determine which units are active and elucidate mechanisms of spatial and temporal transcriptional control. We have detected four variants among rRNA genes of Arabidopsis thaliana by analysis of the 3' external spacer region. Surprisingly, the most abundant variant, representing ~50% of the genes, is not expressed in wild-type plants but is transcribed in lines mutated in one of the two nucleolin genes. Analysis of Arabidopsis ecotypes indicated considerable variability in numbers and presence/absence of variants, although more closely-related ecotypes show similar profiles. Sequence analysis showed that one of the variants is not only unexpectedly located within a 5S RNA gene cluster in the pericentromeric region of chromosome 3 but is also potentially highly expressed in wild-type plants. We present a model to explain how transcription of this this unusual variant, maintained in an active state by nucleolin binding, could be involved in control of expression of the major variant, while absence of nucleolin leads to silencing of the minor variant and consequent expression of the major variant.